In general terms a biofilm is a collection, or community, of microorganisms surrounded by a matrix of extracellular polymers (also known in the art as a glycocalyx). These extracellular polymers are typically polysaccharides, notably polysaccharides produced by the organisms themselves, but they can contain other biopolymers as well. A biofilm will typically be attached to a surface, which may be inert or living, but it has also been observed that biofilms may form from microorganisms attached to each other or at any interface. Generally, therefore, a biofilm is characterised as a highly organised multicellular community of microorganisms encased in, or surrounded by, an extracellular polymer matrix, generally a polysaccharide matrix, and generally in close association with a surface or interface. Such a mode of growth is protective to the microorganisms, and renders them difficult to remove or eradicate (for example, as discussed further below, recalcitrant or resistant to anti-microbial agents or host defence or clearance mechanisms). It is believed, according to the present invention, that alginate oligomers may interact with the polymer matrix of the biofilm, and thus weaken the biofilm. As discussed further below, biofilms cause significant commercial, industrial and medical problems, in terms of infections. contamination, fouling and spoilage etc, and thus the present invention provides a significant advantage in enabling or facilitating the combating of such biofilms, including both reducing or preventing their formation, and rendering them more susceptible to removal or reduction, e.g. more susceptible to the effect of anti-microbial agents (including disinfectants or antibiotics) or indeed in the case of an infection, to the immune response of the infected host. The efficacy of anti-microbial agents, both therapeutic and non-therapeutic and including particularly antibiotics, may thus be enhanced.
Biofilms are found ubiquitously on a wide variety of surfaces or interfaces (e.g. water/solid and water/gas (for example water/air) interfaces) if conditions conducive to microbial colonisation exist. Basically a biofilm will form wherever there are microorganisms and an interface or surface, particularly a surface exposed to water or moisture and biofilms are now recognised as the, natural state of microbial growth on such surfaces or interfaces. In basic terms, as noted above, a biofilm is the complex and organised arrangement of microbial colonies on a surface, or at an interface, which may occur particularly in the presence of water or moisture. The organisation of these colonies results from the ability of microorganisms to produce an organised extracellular matrix in which the cells are “embedded”. This matrix is formed from biopolymers produced by the microorganisms with polysaccharides typically the predominant polymer.
The microorganisms in a biofilm community display properties at the cellular level (phenotype) that are not shared by their planktonic (free-floating) equivalents. In fact, it is believed that microorganisms in a biofilm are profoundly different from planktonic free-floating cells. Further differences can be also be observed at the community level and are attributed to the effects of the extracellular matrix. Perhaps most notable is the commonly observed phenomenon that microorganisms in a biofilm environment do not display the same susceptibilities to anti-microbial agents, e.g. antibiotics, anti-fungals and microbicides, and host immune defences or clearance mechanisms. It is thought that this resistance is due to the barrier effect of the extracellular matrix and/or a phenotypic change in the microbes themselves. For instance, once biofilms form, antibodies no longer attach to the microorganisms (e.g. bacteria) within the biofilm. Experiments have shown antibodies thickly crusted on the outside of biofilm, but not within the biofilm itself. Studies on white blood cell activity against biofilms have demonstrated similar findings. Toxin production might also different between a planktonic microbe and its equivalent residing in a biofilm colony suggesting phenotypic changes in the microbes. It is also believed that microorganisms in biofilms may grow more slowly, and as a result take up anti-microbial agents more slowly.
Biofilms form readily on aquatic environmental surfaces and an established microbial colony on any surface exposed to water (any “wet” surface) will almost certainly exist as a biofilm structure. Furthermore it is now becoming evident and increasingly documented that biofilms may also form in the case of microbial infections i.e. within or on an infected host. Thus biofilm formation may also occur on a “physiological” or “biological” surface, that is on an animate or biotic surface, or a surface on or in an infected host organism (e.g. a human or non-human animal subject), for example on an internal or external body or tissue surface. Such biofilm formation (or infection) on body tissues is increasingly believed to contribute to various infective diseases, including for example native valve endocarditis (mitral, aortic, tricupsid, pulmonic heart valves), acute otitis media (middle ear), chronic bacterial prostatitis (prostate), cystic fibrosis (lungs), pneumonia (respiratory tract), periodontitis (tissues supporting the teeth, e.g. gingiva, periodontal ligament, alvelor bone). Of course, both of these biofilm niches are present when medical devices are implanted and the formation of biofilm on such implanted (“in-dwelling”) devices can lead to clinical problems with infection at such sites, such as prosthetic valve endocarditis and device-related infection, for example with intrauterine devices, contact lenses, prostheses (e.g. prosthetic joints) and at catheterisation sites, for example with central venous or urinary catheters.
A significant problem and risk with such biofilm infections is that microorganisms (or more particularly microcolonies) may break off or detach from the biofilm, and enter other tissues, including significantly the circulation. Such circulating biofilm-derived microorganisms can cause further infections and lead to significant clinical problems, particularly as the detached circulating microorganisms may have all the resistance characteristics of the parent community.
A biofilm infection typically develops gradually and may be slow to produce overt symptoms. Once established, however, biofilms are as noted above difficult to clear and a biofilm infection will typically be persistent, and rarely resolved by host defence or immune mechanisms, even in individuals with healthy innate and adaptive immune responses. Active host responses may indeed be detrimental, for example cell-mediated immunity (e.g. invading neutrophils) may cause collateral damage to neighbouring healthy host tissue. Biofilm infections respond only transiently to antibiotic therapy. Thus, whilst planktonic microbial cells may be cleared by antibodies or phagocytes, and are susceptible to anti-microbials, the microorganisms in biofilms tend to be resistant to antibodies, phagocytes and anti-microbials. Phagocytes are attracted to the biofilm, but phagocytosis is frustrated. Phagocytic enzymes are nonetheless released and may damage tissue around the biofilm. Planktonic bacteria may be released from the biofilm and such release may cause dissemination and acute infection in neighbouring tissue.
Body or tissue surfaces which are dead or damaged (e.g. necrotic or inflamed) are particularly susceptible to biofilm infection. Wounds are susceptible to infection and biofilm formation can occur in wounds that do not heal in a short amount of time. Wounds are an ideal environment for the formation of biofilms due to their susceptibility to bacterial colonisation and the availability of substrate and surface for biofilm attachment. Problematically, infection of a wound often delays healing further and thus renders that wound more susceptible to biofilm formation and established infection. Wounds in which healing is delayed (so called chronic wounds) represent sites of particular concern with respect to biofilm formation. A chronic wound is in an inflammatory state, with elevated levels of pro-inflammatory cytokines. The effect of these cytokines is to produce a swarming of the area with immune cells (neutrophils and macrophages). If this defence system is in any way delayed (as in chronic wounds), bacteria or other microorganisms have time to attach to the surface and enter the biofilm mode of growth. Evidence is increasingly growing that both chronic and acute wounds may be sites of biofilm infection, with evidence of diverse microbial communities or populations in wounds, particularly chronic wounds, including anaerobic bacteria within chronic wounds. Chronic wound infections share two important attributes with other biofilm infections: persistent infection that is not cleared by the host immune system even in individuals with healthy innate and adaptive immune reactions, and increased resistance to systemic and topical antimicrobial agents. Accordingly, biofilm based infection is very difficult to treat and biofilm contamination is very difficult to eradicate. Frequent debridement is one of the most clinically effective treatments to help heal chronic wounds. This is an effective treatment, in part, because it physically removes the biofilm from the wound. This is similar in principle to resolving infections from biofilm-colonized in-dwelling medical devices (e.g. catheters)—where antibiotic therapy is ineffective the most effective approach is to remove or replace the biofilm-infected device.
Chronic wounds are a major health problem throughout the world and represent a significant drain on clinical resources. Three principle types of chronic wound are diabetic foot ulcers, venous leg ulcers and pressure ulcers, although other wounds, including surgical wounds, may become chronic. The care of such wound imposes enormous material and patient costs, and hence an effective anti-biofilm treatment, or indeed any treatment which assisted in or facilitated the treatment of biofilms, and thus accelerated or facilitated wound healing, would be of very significant impact.
More generally, given the widespread occurrence of biofilms and the medical, environmental, industrial or other commercial problems they cause, any means of improving or enabling the combating of biofilms would be very important, both clinically and commercially.